title: Enhancement of autophagy by simvastatin through inhibition of Rac1-mTOR signaling pathway in coronary arterial myocytes.
abstract: OBJECTIVE
  In addition to their action of lowering blood cholesterol levels, statins modulate biological characteristics and functions of arterial myocytes such as viability, proliferation, apoptosis, survival and contraction. The present study tested whether simvastatin, as a prototype statin, enhances autophagy in coronary arterial myocytes (CAMs) to thereby exert their beneficial effects in atherosclerosis.
  
  
  RESULTS
  Using flow cytometry, we demonstrated that simvastatin significantly increased the autophagsome formation in CAMs. Western blot analysis confirmed that simvastatin significantly increased protein expression of typical autophagy markers LC3B and Beclin1 in these CAMs. Confocal microscopy further demonstrated that simvastatin increased fusion of autophagosomes with lysosomes, which was blocked by autophagy inhibitor 3-methyladenine or silencing of Atg7 genes. Simvastatin reduced mammalian target of rapamycin (mTOR) activity, which was reversed by Rac1-GTPase overexpression and the mTOR agonist phosphatidic acid. Moreover, both Rac1-GTPase overexpression and activation of mTOR by phosphatidic acid drastically blocked simvastatin-induced autophagosome formation in CAMs. Interestingly, simvastatin increased protein expression of a contractile phenotype marker calponin in CAMs, which was blocked by autophagy inhibitor 3-methyladenine. Simvastatin markedly reduced proliferation of CAMs under both control and proatherogenic stimulation. However, this inhibitory effect of simvastatin on CAM proliferation was blocked by by autophagy inhibitor 3-methyladenine or silencing of Atg7 genes. Lastly, animal experiments demonstrated that simvastatin increased protein expression of LC3B and calponin in mouse coronary arteries.
  
  
  CONCLUSIONS
  Our results indicate that simvastatin inhibits the Rac1-mTOR pathway and thereby increases autophagy in CAMs which may stabilize CAMs in the contractile phenotype to prevent proliferation and growth of these cells.
authors.affiliation: Department of Pharmacology & Toxicology, Medical College of Virginia Campus, Virginia Commonwealth University, Richmond, VA 23298, USA.
authors.forename: Yu-Miao
authors.initials: YM
authors.lastname: Wei
authors.forename: Xiang
authors.initials: X
authors.lastname: Li
authors.forename: Ming
authors.initials: M
authors.lastname: Xu
authors.forename: Justine M
authors.initials: JM
authors.lastname: Abais
authors.forename: Yang
authors.initials: Y
authors.lastname: Chen
authors.forename: Christopher R
authors.initials: CR
authors.lastname: Riebling
authors.forename: Krishna M
authors.initials: KM
authors.lastname: Boini
authors.forename: Pin-Lan
authors.initials: PL
authors.lastname: Li
authors.forename: Yang
authors.initials: Y
authors.lastname: Zhang
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001343:Autophagy / Q000187:drug effects*; D000071193:Autophagy-Related Protein 7; D002135:Calcium-Binding Proteins / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology*; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000494:pharmacology*; D008247:Lysosomes / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008840:Microfilament Proteins / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D032342:Muscle Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D010641:Phenotype; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction; D019821:Simvastatin / Q000494:pharmacology*; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000096985:Calponins
chemical_list: D000225:Adenine; D000071193:Autophagy-Related Protein 7; D002135:Calcium-Binding Proteins; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D008840:Microfilament Proteins; D008869:Microtubule-Associated Proteins; D034741:RNA, Small Interfering; D019821:Simvastatin; D020123:Sirolimus; D058570:TOR Serine-Threonine Kinases; D020830:rac1 GTP-Binding Protein; D000096985:Calponins; C502661:Atg7 protein, mouse; C480860:Map1lc3b protein, mouse; C025946:3-methyladenine